Petrich de Marquesini Liliana G, Moustakas Antonis K, Thomas Ian J, Wen Li, Papadopoulos George K, Wong F Susan
Department of Cellular and Molecular Medicine, School of Medical Sciences, University of Bristol, Bristol, UK.
Eur J Immunol. 2008 Jan;38(1):240-9. doi: 10.1002/eji.200737762.
Insulin-reactive CD8 T cells are amongst the earliest islet-infiltrating CD8 T cells in NOD mice. Cloned insulin B15-23-reactive cells (designated G9C8), restricted by H-2K(d), are highly diabetogenic. We used altered peptide ligands (APL) substituted at TCR contact sites, positions (p)6 and 8, to investigate G9C8 T cell function and correlated this with structure. Cytotoxicity and IFN-gamma production assays revealed that p6G and p8R could not be replaced by any naturally occurring amino acid without abrogating recognition and functional response by the G9C8 clone. When tested for antagonist activity with APL differing from the native peptide at either of these positions, the peptide variants, G6H and R8L showed the capacity to reduce the agonist response to the native peptide. The antagonist activity in cytotoxicity and IFN-gamma production assays can be correlated with conformational changes induced by different structures of the MHC-peptide complexes, shown by molecular modeling. We conclude that p6 and p8 of the insulin B15-23 peptide are very important for TCR stimulation of this clone and no substitutions are tolerated at these positions in the peptide. This is important in considering the therapeutic use of peptides as APL that encompass both CD4 and CD8 epitopes of insulin.
胰岛素反应性CD8 T细胞是NOD小鼠中最早浸润胰岛的CD8 T细胞之一。克隆的胰岛素B15 - 23反应性细胞（命名为G9C8）受H - 2K(d)限制，具有高度致糖尿病性。我们使用在TCR接触位点（第6和8位）进行取代的改变肽配体（APL）来研究G9C8 T细胞功能，并将其与结构相关联。细胞毒性和IFN - γ产生测定表明，第6位甘氨酸（p6G）和第8位精氨酸（p8R）不能被任何天然存在的氨基酸取代，否则会消除G9C8克隆的识别和功能反应。当用在这些位置之一与天然肽不同的APL测试其拮抗活性时，肽变体G6H和R8L显示出降低对天然肽激动剂反应的能力。细胞毒性和IFN - γ产生测定中的拮抗活性可与MHC - 肽复合物不同结构诱导的构象变化相关联，分子建模显示了这一点。我们得出结论，胰岛素B15 - 23肽的第6位和第8位对于该克隆的TCR刺激非常重要，肽在这些位置不容许有任何取代。这对于考虑将包含胰岛素CD4和CD8表位的肽作为APL用于治疗具有重要意义。
